LOCUS       sequence-171004-        7154 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor vector for 3' FLAG tag of human HLF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7154)
  AUTHORS   Savic D, Partridge EC, Newberry KM, Smith SB, Meadows SK, Roberts 
            BS, Mackiewicz M, Mendenhall EM, Myers RM
  TITLE     CETCh-seq: CRISPR epitope tagging ChIP-seq of DNA-binding proteins.
  JOURNAL   Genome Res. 2015 Sep 9.
  PUBMED    26355004
REFERENCE   2  (bases 1 to 7154)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Genome Res.
            2015 Sep 9."
COMMENT     Sequence Label: pFETCh_HLF
FEATURES             Location/Qualifiers
     source          1..7154
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      33..621
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     522..541
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     775..792
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1092..1113
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1128..1133,1134..1151,1152..1158)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1166..1182
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     1171..1193
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1190..1206
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1190..1206
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        1383..1585
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1536..1556
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1582..1606
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1682..2812
                     /codon_start=1
                     /product="herpes simplex virus thymidine kinase"
                     /label=HSV TK
                     /note="confers sensitivity to nucleoside analogs such as 
                     acyclovir or ganciclovir"
                     /translation="MASYPCHQHASAFDQAARSRGHSNRRTALRPRRQQEATEVRLEQK
                     MPTLLRVYIDGPHGMGKTTTTQLLVALGSRDDIVYVPEPMTYWQVLGASETIANIYTTQ
                     HRLDQGEISAGDAAVVMTSAQITMGMPYAVTDAVLAPHIGGEAGSSHAPPPALTLIFDR
                     HPIAALLCYPAARYLMGSMTPQAVLAFVALIPPTLPGTNIVLGALPEDRHIDRLAKRQR
                     PGERLDLAMLAAIRRVYGLLANTVRYLQGGGSWREDWGQLSGTAVPPQGAEPQSNAGPR
                     PHIGDTLFTLFRAPELLAPNGDLYNVFAWALDVLAKRLRPMHVFILDYDQSPAGCRDAL
                     LQLTSGMVQTHVTTPGSIPTICDLARTFAREMGEAN"
     CDS             3938..3961
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3965..3988
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3992..4015
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             4031..4087
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             4100..4894
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4154..4173)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4764..4783
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5757..5773)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5766..5788)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(5992..6807)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNILQFHLMLDEFF"
     primer_bind     6715..6734
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     6799..6818
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
       61 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
      121 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
      181 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
      241 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
      301 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
      361 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
      421 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
      481 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
      541 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
      601 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
      661 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
      721 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
      781 tcagtgagcg aggaagcgga agaagctcgc acattcagca gcgtttttca gcgcgttttc
      841 gatcaacgtt tcaatgttgg tatcaacacc aggtttaact ttgaacttat cggcactgac
      901 ggttactgat tttgaacttt tgctttgcca cggaacggtc tgcgttgtcg ggaagatgcg
      961 tgatctgatc cttcaactca gcaaaagttc ccaatacgca aaccgcctct ccccgcgcgt
     1021 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     1081 cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg
     1141 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
     1201 tatgaccatg attacgccaa gcttgcatgc ctgcaggtcg actctagagg atccccgggt
     1261 accgagctcg aattagcttg catgcctgca ggtcaattcc ctggcattat gcccagtaca
     1321 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
     1381 tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat
     1441 ttcaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga
     1501 ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg
     1561 gtgggaggtc tatataagca gagctcgttt agtgaaccgt cagatcgcct ggagacgcca
     1621 tccacgctgt tttgacctcc ataggagaca ccgggaccga tccagcctgg ggatcccgac
     1681 catggcttcg tacccctgcc atcaacacgc gtctgcgttc gaccaggctg cgcgttctcg
     1741 cggccatagc aaccgacgta cggcgttgcg ccctcgccgg cagcaagaag ccacggaagt
     1801 ccgcctggag cagaaaatgc ccacgctact gcgggtttat atagacggtc ctcacgggat
     1861 ggggaaaacc accaccacgc aactgctggt ggccctgggt tcgcgcgacg atatcgtcta
     1921 cgtacccgag ccgatgactt actggcaggt gctgggggct tccgagacaa tcgcgaacat
     1981 ctacaccaca caacaccgcc tcgaccaggg tgagatatcg gccggggacg cggcggtggt
     2041 aatgacaagc gcccagataa caatgggcat gccttatgcc gtgaccgacg ccgttctggc
     2101 tcctcatatc gggggggagg ctgggagctc acatgccccg cccccggccc tcaccctcat
     2161 cttcgaccgc catcccatcg ccgccctcct gtgctacccg gccgcgcgat accttatggg
     2221 cagcatgacc ccccaggccg tgctggcgtt cgtggccctc atcccgccga ccttgcccgg
     2281 cacaaacatc gtgttggggg cccttccgga ggacagacac atcgaccgcc tggccaaacg
     2341 ccagcgcccc ggcgagcggc ttgacctggc tatgctggcc gcgattcgcc gcgtttacgg
     2401 gctgcttgcc aatacggtgc ggtatctgca gggcggcggg tcgtggcggg aggattgggg
     2461 acagctttcg gggacggccg tgccgcccca gggtgccgag ccccagagca acgcgggccc
     2521 acgaccccat atcggggaca cgttatttac cctgtttcgg gcccccgagt tgctggcccc
     2581 caacggcgac ctgtacaacg tgtttgcctg ggccttggac gtcttggcca aacgcctccg
     2641 tcccatgcac gtctttatcc tggattacga ccaatcgccc gccggctgcc gggacgccct
     2701 gctgcaactt acctccggga tggtccagac ccacgtcacc acccccggct ccataccgac
     2761 gatctgcgac ctggcgcgca cgtttgcccg ggagatgggg gaggctaact gaggatccca
     2821 cgtcactatt gtatactcta tattatactc tatgttatac tctgtaatcc tactcaataa
     2881 acgtgtcacg cctgtgaaac cgtactaagt ctcccgtgta ttcttatcac catcaggtga
     2941 catcctcgcc caggctgtca atcatgccgg tatcgattcc agtagcaccg gccccacgct
     3001 gacaacccac tcttgcagcg ttagcagcgc ccctcttaac aagccgaccc ccaccagcgt
     3061 cgcggttact aacactcctc tccccgacct gcagcccagc tcctatgcat agtttttcct
     3121 cattttttct actgcacata atgctgcagt gaacagttta tgcttgtgtg tacatgcaca
     3181 tgtatggaga ttgctacagg acaaaatcct ggaagtggag tgctagatga atagtatatc
     3241 catttaaata gttgatagat attgccaaat tgccttctaa aatttttgta ccaactcata
     3301 cccaaaaaag catatgagtg cccttgaagg aaaagggttc ttaaaatgtt gagcagggag
     3361 agtaaagaaa gttgaaatgt aatttaaggg tgaggtgagc cacctgcttt catccagcct
     3421 ctgatctctg cttcactgag cacgcctctg ctgggcaaag agtaggagcc tgttgggaca
     3481 gatcggtggg tccttgctcc aagaaggaag tgtggctcat gggccacact ctcagacatg
     3541 cagaaactgt aaaggaagga gacacaagct aagaaacccg ggctggcacc tctgcacaat
     3601 cctggagcct gcccgtgccc tggctggcac tgggctttgc tgaaatctaa tatcttgttt
     3661 ctttttccct tctgtaacta atgagcagga tgacaagtac tgggcaaggc gcagaaagaa
     3721 caacatggca gccaagcgct cccgcgacgc ccggaggctg aaagagaacc agatcgccat
     3781 ccgggcctcg ttcctggaga aggagaactc ggccctccgc caggaggtgg ctgacttgag
     3841 gaaggagctg ggcaaatgca agaacatact tgccaagtat gaggccaggc acgggcccct
     3901 ggggagcgga ggaggttccg gtggaggtgg ttctggagat tacaaggatg acgacgataa
     3961 gggcgattac aaggatgacg acgataaggg agattacaag gatgacgacg ataaggtttc
     4021 aggaagcgga gctactaact tcagcctgct gaagcaggct ggagacgtgg aggagaaccc
     4081 tggacctgga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
     4141 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
     4201 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
     4261 cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg
     4321 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
     4381 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
     4441 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
     4501 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
     4561 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
     4621 caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
     4681 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
     4741 ggccgaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     4801 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     4861 cgccttctat cgccttcttg acgagttctt ctgattcgaa catcggctgg ctttggaata
     4921 gatggacagt ttgtttcctg tctgatagca ccacacgcaa accaaccttt ctgacatcag
     4981 cactttacca gaggcataaa cacaactgac tcccattttg gtgtgcatct gtgtgtgtgt
     5041 gcgtgtatat gtgcttgtgc tcatgtgtgt ggtcagcggt atgtgcgtgt gcgtgttcct
     5101 ttgctcttgc cattttaagg tagccctctc atcgtctttt agttccaaca aagaaaggtg
     5161 ccatgtcttt actagactga ggagccctct cgcgggtctc ccatcccctc cctccttcac
     5221 tcctgcctcc tcagctttgc ttcatgttcg agcttaccta ctcttccagg actctctgct
     5281 tggattcact aaaaagggcc ctggtaaaat agtggatctc agtttttaag agtacaagct
     5341 cttgtttctg tttagtccgt aagttaccat gctaatgagg tgcacacaat aacttagcac
     5401 tactccgcag ctctagtcct ttataagttg ctttcctctt actttcagtt ttggtgataa
     5461 tcgtcttcaa attaaagtgc tgtttagatt tattagatcc catatttact tactgctatc
     5521 tactaagttt ccttttaatt ctaccaaccc cagataagta agagtactat taatagaaca
     5581 cagagtgtgt ttttgcactg tctgtaccta aagcaataat cctattgtac gctagagcat
     5641 gctgcctgag tattactagt ggacgtagga tattttccct acctaagaat ttcactgtct
     5701 tttacttgga aagtcctctc cactgactgt agcctccaat tcactggccg tcgttcactg
     5761 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     5821 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     5881 tcccaacagt tgcgcagcct gaatggcgaa tgcgatttat tcaacaaagc cgccgtcccg
     5941 tcaagtcagc gtaatgctct gccagtgtta caaccaatta accaattctg attagaaaaa
     6001 ctcatcgagc atcaaatgaa actgcaatat attcatatca ggattatcaa taccatattt
     6061 ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc ataggatggc
     6121 aagatcctgg tatcggtctg cgattccgac tcgtccaaca tcaatacaac ctattaattt
     6181 cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga ctgaatccgg
     6241 tgagaatggc aaaagcttat gcatttcttt ccagacttgt tcaacaggcc agccattacg
     6301 ctcgtcatca aaatcactcg catcaaccaa accgttattc attcgtgatt gcgcctgagc
     6361 gagacgaaat acgcgatcgc tgttaaaagg acaattacaa acaggaatcg aatgcaaccg
     6421 gcgcaggaac actgccagcg catcaacaat attttcacct gaatcaggat attcttctaa
     6481 tacctggaat gctgttttcc cggggatcgc agtggtgagt aaccatgcat catcaggagt
     6541 acggataaaa tgcttgatgg tcggaagagg cataaattcc gtcagccagt ttagtctgac
     6601 catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa acaactctgg
     6661 cgcatcgggc ttcccataca atcgatagat tgtcgcacct gattgcccga cattatcgcg
     6721 agcccattta tacccatata aatcagcatc catgttggaa tttaatcgcg gcttcgagca
     6781 agacgtttcc cgttgaatat ggctcataac accccttgta ttactgttta tgtaagcaga
     6841 cagttttatt gttcatgatg atatattttt atcttgtgca atgtaacatc agagattttg
     6901 agacacaacg tggctttgtt gaataaatcg aacttttgct gagttgaagg atcagatcac
     6961 gcatcttccc gacaacgcag accgttccgt ggcaaagcaa aagttcaaaa tcaccaactg
     7021 gtccacctac aacaaagctc tcatcaaccg tggctccctc actttctggc tggatgatgg
     7081 ggcgattcag gcctggtatg agtcagcaac accttcttca cgaggcagac ctctcgacgg
     7141 agttccactg agcg
//
